Broad Spectrum Beta-Lactamase Inhibitors

ABSTRACT

Broad spectrum beta-lactamase inhibitors. Certain inhibitors also exhibit potent antibiotic activity in addition to beta-lactamase inhibition. Compounds of the invention are designed such that on cleavage of the beta-lactam ring reactive moieties are generated which can inactivate beta-lactamase. Also provided are methods of making beta-lactamase inhibitors and beta-lactam antibiotics exhibiting such inhibition. Additionally provided are pharmaceutical compositions for treatment or prevention of bacterial infections and methods of treatment of such infections.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.15/247,536, filed Aug. 25, 2016, which is a division of U.S. applicationSer. No. 14/491,806, filed Sep. 19, 2014, now U.S. Pat. No. 9,453,032,which in turn is a continuation of U.S. application Ser. No. 12/248,760,filed Oct. 9, 2008, now U.S. Pat. No. 8,883,772, which claims thebenefit of and priority to U.S. Provisional Application Nos. 60/997,898and 60/997,941, both filed Oct. 9, 2007. Each of these applications isincorporated by reference herein in its entirety.

BACKGROUND OF THE INVENTION

The present invention relates to beta-lactamase inhibitor compounds,their production and use.

The invention and use of antibiotics to cure infectious diseases causedby bacteria is one of the milestones of modem medical and scientifictechnology. The beta-lactam class of antibiotics has been and continuesto be one of the most important. Antibiotic resistance has become amajor problem worldwide. One of the most important resistance mechanismsto beta-lactam antibiotics is the bacterial production ofbeta-lactamases, enzymes that inactivate beta-lactam antibiotics bycatalyzing the hydrolysis of the lactam ring rendering the antibioticsineffective towards binding of their target, penicillin binding protein.

Previous attempts to circumvent inactivation by beta-lactamases havebeen to alter beta-lactam compounds by functionalizing them with variousorganic groups conferring resistance to beta-lactam hydrolysis whilemaintaining antimicrobial potency. However, evolution of beta-lactamaseshas kept pace and there is now a beta-lactamase that is able toinactivate every known clinically available beta-lactam antibiotic; over500 beta-lactamases have been documented.

Broadly defined by mechanism there are two fundamental classes ofbeta-lactamases, serine hydrolases and metallo-hydrolases. The enzymescan be further classified by subdividing them into groups according totheir spectrum of activity towards beta-lactam compounds. The serinehydrolases are sub-classified into Bush Class A which are thepenicillinases. Class C enzymes refer to the cephalosporinases. WhileClass D enzymes are the broad spectrum or extended spectrumbeta-lactamases (ESBL). Bush Class B beta-lactamases refer to themetallo-enzymes that require one or two Zn2+ ions for activity andlikewise show a broad spectrum of activity towards beta-lactamantibiotics. Another strategy has been to develop and use inhibitors ofbeta-lactamases. Three compounds are currently in clinical use,clavulanic acid, sulbactam and tazobactam.

These compounds irreversibly inhibit Class A penicillinases. Drawbacksof the known inhibitors are that they possess little intrinsicantimicrobial activity and therefore must be used in combination withbeta-lactam antibiotics. The second shortcoming is that they are notclinically effective at inhibiting Classes B, C, and D enzymes which areincreasingly important.

Thus, there is a significant need in the art for potent beta-lactamantibiotic compositions which demonstrate the additional functionalityof potent beta-lactamase inhibition while maintaining antimicrobialpotency.

SUMMARY OF THE INVENTION

The invention relates to compounds which are beta-lactamase inhibitorsand particularly relates to beta-lactam antibiotics that also exhibitinhibition of beta-lactamase. The invention is further directed tomethods of making such compounds and methods of using such compounds forinhibition of microbial growth.

In an embodiment, the invention provides compounds of formula I:

and pharmacologically acceptable salts thereof wherein:

R is a pharmaceutically acceptable functional group including, anacylamino group, and pharmaceutically acceptable salts thereof;

R¹, R², R³, R⁴ and R⁵ are selected from hydrogen or a wide range oforganic groups;

n is an integer ranging from 1-5 and is preferably 1;

- - - Z - - - - is a linker between the two indicated atoms and ispresent or absent, when it is absent y is 2, when it is present y is 1;

- - - Z - - - is a one or two atom linker which forms a 5 or 6 memberring Z can be two carbon atoms, a carbon and a sulfur atom, a carbon anda nitrogen, or a carbon and an oxygen, where any remaining valences aresatisfied by substitution of atoms with hydrogen or organicsubstituents, e.g., alkyl groups;

M can be cis or trans with respect to R¹; and

M most generally represents a chemical species which is in conjugationwith the nitrogen of the core beta-lactam ring system of the compound,such that one or more reactive species, e.g., electrophilic ornucleophilic sites are generated on modification of M which is initiatedby cleavage of the beta-lactam ring.

In specific embodiments, R is an acylamino group of a known beta-lactamantibiotic. A wide variety of beta-lactam antibiotics is known in theart. Acylamino groups of representative known beta-lactam antibioticsare described herein after.

Certain M groups of this invention contain good chemical leaving groupswhich are caused to cleave from the M group by beta-lactam ringcleavage. Beta-lactam cleavage is initiated by attack of abeta-lactamase enzyme on the compound. The reactive groups generated inM on cleavage of the beta-lactam ring are available for reaction withthe beta-lactamase and function to inhibit the activity of thebeta-lactamase.

Beta-lactam ring systems of the compounds of this invention includethose of cephems, cephamycins, carbacephems, penems, and monobactams.

The invention provide compounds of Formula I as generally describedabove and as more specifically described hereinafter, for use asbeta-lactamase inhibitors and beta-lactam antibiotics. Compounds of thisinvention can exhibit one or both of these functions and as such areuseful in a variety of therapeutic (human and veterinary) applicationsfor treatment of microbial infections and complications thereof. Thecompounds of this invention are particularly useful for treatment ofinfections of microorganisms, particularly bacteria which are known toexhibit resistance to one or more beta-lactam antibiotics. The compoundsof this invention are useful for inhibition of growth of microorganismsincluding bacteria in vivo or in vitro applications. Beta-lactaminhibitors of this invention may be combined with beta-lactamantibiotics to provide for inhibition of beta-lactamases in vivo or invivo applications.

Compound of this invention including M groups as described above and inwhich R is not an aminoacyl group and those in which R is A-CO—NH, whereA is an unsubstituted alkyl or aryl group (e.g., phenyl group) areuseful as intermediates in the synthesis of beta-lactam inhibitors andbeta-lactam antibiotics which exhibit beta-lactamase inhibition and inwhich the aminoacyl group is that of a known beta-lactam antibiotic. Abeta-lactam inhibitor which does not exhibit antibiotic activity or inwhich it is desired to improve antibiotic activity can be prepared fromM group containing compound of this invention by replacing the R groupwith a selected aminoacyl which is found in a beta-lactam antibioticwhich is known in the art. Thus, this invention provides a method formaking improved beta-lactam antibiotics which exhibit beta-lactamaseinhibition in addition to antibiotic activity.

The invention is further related to pharmaceutical compositionscomprising one or more compounds of this invention of formula I andother formulas described herein after.

The invention is also related to method of treatment of infections andrelated disorders, diseases or complications by administering atherapeutically effective amount or combined amount of one or morecompounds of the invention optionally in combination with atherapeutically effective amount or a combined amount of one or moreknown beta-lactam antibiotics.

The invention is further related to a method of inhibiting the growth ofmicroorganisms, particularly bacteria, by contacting the microorganismin vivo or in vitro with an amount of one or more of the compounds ofthis invention, optionally in combination with a known beta-lactamantibiotic, particularly an antibiotic that has been used in the past oris currently used for therapeutic applications (in humans or animals).

The invention also relates to method for making medicaments comprisingone or more compounds of this invention, particularly for treatment ofinfections and related disorders, diseases or complications thereof.

The invention is further described and illustrated in the followingdetailed description, examples and drawings which, however, are notintended to be limiting. Additional, aspects and embodiments of theinvention will be apparent on review of the specification as a whole.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph of time dependent Inhibition of a beta-lactamase by3-vinylcyclopropane-7-(2-Phenylacetamido)-3-Cephem-4-carboxylic acid(IX).

FIG. 2 is a graph of time dependent Inhibition of a beta-lactamase by3-(1-bromomethyl-4-vinylbenzene)-7-(2-phenylacetamido)-3-Cephem-4-carboxylicacid (XI).

FIGS. 3-1 through 3-2 illustrate exemplary acyl groups of compounds ofthe formulas herein.

FIGS. 4-1 through 4-2 illustrate additional exemplary structures ofcompounds of the invention.

FIGS. 5-1 through 5-6 illustrate additional exemplary M groups of thecompounds of the invention.

FIG. 6 illustrates preferred stereochemistry of various core beta-lactamring structures of the formulas of this invention.

DETAILED DESCRIPTION OF THE INVENTION

The invention relates to methods for making improved beta-lactamantibiotic which exhibit inhibition of one or more beta-lactamases inaddition to antibacterial activity. The invention also relates tocertain beta-lactam compounds exhibiting inhibition of one or morebeta-lactamases. The invention further relates to certain beta-lactamcompounds exhibiting beta-lactamase inhibition and antibiotic activity.In specific embodiments, compounds of the invention inhibit one or morebeta-lactamases in addition to the Class A penicillinases. In specificembodiments, compounds of the invention inhibit beta-lactamases otherthan the Class A penicillinases. In specific embodiments, compounds ofthe invention exhibit inhibition of one or more Class B, C or Dbeta-lactamases. In specific embodiments, compounds of the inventionexhibit broad spectrum inhibition of one or more beta-lactamases ofdifferent classes. In specific embodiments, compounds of the inventionexhibit irreversible inhibition of one or more beta-lactamases.

In specific embodiments the invention relates to beta-lactam compoundsof formula:

and pharmacologically acceptable salts thereof wherein:

- - - - Z - - - is present or absent and represents —O—(CH₂)x-,—CH₂—(CH₂)x-, —NR′—(CH₂)x-, —S—(CH₂)x-, —SO—(CH₂)x-, or —SO₂—(CH₂)x-,where x is 0 or 1 and R′ is hydrogen or C1-C6 alkyl, when - - - -Z - - - is present y is 1 and when it is absent y is 2;

n is an integer from 1-5;

R¹ and R², independently, are selected from the group consisting ofhydrogen, halogen, (C1-C6) alkyl, (C1-C6) alkoxy and (C1-C6) thioalkoxy(—S-alkyl);

Y is O—C+ or OR³ where R³ is hydrogen, or an optionally substitutedalkyl or aryl group and C+ is a pharmacologically acceptable cation;

R⁴ is hydrogen, (C1-C6) alkyl, OR′, where R′ is hydrogen or (C1-C6)alkyl;

R⁵ is hydrogen, (C1-C6) alkyl;

R is selected from hydrogen, (C1-C6) alkyl group, (C2-C6) alkenyl group,(C2-C6) alkynyl group, (C7-C19) aralkyl group, a 3-7-member-ring cyclichydrocarbon group, a 3-7 member heterocyclic group, a (C6-C10) aromaticgroup, a 6-10 member heterocyclic aromatic group, a —CO—R″, —CO₂R″,—CO—N(R′)₂; —N(R″)₂, —NRCO₂—R″ wherein each R″ is independently selectedfrom hydrogen, (C1-C6) alkyl group, (C2-C6) alkenyl group, (C2-C6)alkynyl group, (C7-C19) aralkyl group, a 3-7-member-ring cyclichydrocarbon group, a 3-7 member heterocyclic group, a (C6-C10) aromaticgroup, a 6-10 member heterocyclic aromatic group, each of said groups isoptionally substituted, and an acylamine group A-CO—NH—; and

M, which may be in the cis or trans position with respect to R¹, isselected from groups P, B, BZ, D, E, F as follows:

where:

W is O or C(R″) and each R″, independently, is selected from the groupconsisting of hydrogen, halogen, (C1-C6) alkyl group, (C2-C6) alkenylgroup, (C2-C6) alkynyl group, (C7-C19) aralkyl group, a 3-7-member-ringcyclic hydrocarbon group, a 3-7 member heterocyclic group, a (C6-C10)aromatic group and a 6-10 member heterocyclic aromatic group whereineach of said groups is optionally substituted; and

R⁶ and R⁷ are independently selected from hydrogen, halogen, (C1-C6)alkyl group, (C2-C6) alkenyl group, (C2-C6) alkynyl group, (C7-C19)aralkyl group, a 3-7-member-ring cyclic hydrocarbon group, a 3-7 memberheterocyclic group, a (C6-C10) aromatic group, a 6-10 memberheterocyclic aromatic group, —COR′—,

—COOR″, —CON(R″)₂, wherein each R″, independently, is selected from thegroup consisting of hydrogen, halogen, (C1-C6) alkyl group, (C2-C6)alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkyl group, a3-7-member-ring cyclic hydrocarbon group, a 3-7 member heterocyclicgroup, a (C6-C10) aromatic group and a 6-10 member heterocyclic aromaticgroup and each of said groups is optionally substituted;

where:

Z² is O, NR¹¹ or S where R¹¹ is selected from the group consisting ofhydrogen, (C1-C6) alkyl group, (C2-C6) alkenyl group, and (C2-C6)alkynyl group, where in each group is optionally substituted;

each R⁸ is independently selected from hydrogen, halogen, (C1-C6) alkylgroup, (C2-C6) alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkylgroup, a 3-7-member-ring cyclic hydrocarbon group, a 3-7 memberheterocyclic group, a (C6-C10) aromatic group and a 6-10 memberheterocyclic aromatic group, a (C1-C6) alkoxy group, a (C1-C6) thioalkylgroup, a —CO—R′, —CO₂R′, —CO—N(R′)₂; —N(R′)₂, —NR—CO—R′, —NRCO₂—R′wherein each R′ is independently selected from hydrogen, (C1-C6) alkylgroup, (C2-C6) alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkylgroup, a 3-7-member-ring cyclic hydrocarbon group, a 3-7 memberheterocyclic group, a (C6-C10) aromatic group and a 6-10 memberheterocyclic aromatic group, each of said groups is optionallysubstituted; and

each R⁹ is independently selected from hydrogen, (C1-C6) alkyl group,(C2-C6) alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkyl group, a3-7-member-ring cyclic hydrocarbon group, a 3-7 member heterocyclicgroup, a (C6-C10) aromatic group and a 6-10 member heterocyclic aromaticgroup wherein each of said groups is optionally substituted;

each R¹⁰ is independently selected from the group consisting ofhydrogen, halogen, (C1-C6) alkyl group, (C2-C6) alkenyl group, (C2-C6)alkynyl group, (C7-C19) aralkyl group, a 3-7-member-ring cyclichydrocarbon group, a 3-7 member heterocyclic group, a (C6-C10) aromaticgroup and a 6-10 member heterocyclic aromatic group, a (C1-C6) alkoxygroup, a (C1-C6) thioalkyl group, a —CO—R′, —CO₂R′, —CO—N(R′)₂; —N(R′)₂,—NR—CO—R′, —NRCO₂—R′ wherein each R′ is independently selected fromhydrogen, (C1-C6) alkyl group, (C2-C6) alkenyl group, (C2-C6) alkynylgroup, (C7-C19) aralkyl group, a 3-7-member-ring cyclic hydrocarbongroup, a 3-7 member heterocyclic group, a (C6-C10) aromatic group, a6-10 member heterocyclic aromatic group wherein each of said groups isoptionally substituted, and a —CH₂—X group;

R¹² is selected from the group consisting of hydrogen, (C1-C6) alkylgroup, (C2-C6) alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkylgroup, a 3-7-member-ring cyclic hydrocarbon group, a 3-7 memberheterocyclic group, a (C6-C10) aromatic group and a 6-10 memberheterocyclic aromatic group, a (C1-C6) alkoxy group, a (C1-C6) thioalkylgroup, a —CO—R′, —CO₂R′, —CO—N(R′)₂; —N(R′)₂, —NR—CO—R′, —NRCO₂—R′wherein each R′ is independently selected from hydrogen, (C1-C6) alkylgroup, (C2-C6) alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkylgroup, a 3-7-member-ring cyclic hydrocarbon group, a 3-7 memberheterocyclic group, a (C6-C10) aromatic group, a 6-10 memberheterocyclic aromatic group, wherein each of said groups is optionallysubstituted, and X;

R¹³ and R¹⁴ are independently selected from the group consisting ofhydrogen, (C1-C6) alkyl group, (C2-C6) alkenyl group, (C2-C6) alkynylgroup, (C7-C19) aralkyl group, a 3-7-member-ring cyclic hydrocarbongroup, a 3-7 member heterocyclic group, a (C6-C10) aromatic group and a6-10 member heterocyclic aromatic group, a (C1-C6) alkoxy group, a(C1-C6) thioalkyl group, a —CO—R′, —CO₂R′, —CO—N(R′)₂; —N(R′)₂,—NR—CO—R′, —NRCO₂—R′ wherein each R′ is independently selected fromhydrogen, (C1-C6) alkyl group, (C2-C6) alkenyl group, (C2-C6) alkynylgroup, (C7-C19) aralkyl group, a 3-7-member-ring cyclic hydrocarbongroup, a 3-7 member heterocyclic group, a (C6-C10) aromatic group, a6-10 member heterocyclic aromatic group wherein each of said groups isoptionally substituted, and X; and

X is a leaving group as defined below;

wherein in structure V at least one of R¹⁰ is a —CH₂—X group, R¹² is Xor both; and in structure VI one of R¹³ or R¹⁴ is X.

In specific embodiments, A-CO— is selected from the group consisting of:

where:

R″ for these structures is selected from hydrogen, (C1-C6) alkyl group,(C2-C6) alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkyl group, a3-7-member-ring cyclic hydrocarbon group, a 3-7 member heterocyclicgroup, a (C6-C10) aromatic group and a 6-10 member heterocyclic aromaticgroup,

R′″ for these structures is selected from the group consisting ofhydrogen, (C1-C6) alkyl group, (C2-C6) alkenyl group, (C2-C6) alkynylgroup, (C7-C19) aralkyl group, a 3-7-member-ring cyclic hydrocarbongroup, a 3-7 member heterocyclic group, a (C6-C10) aromatic group and a6-10 member heterocyclic aromatic group, a —CO—R²⁹, —CO₂R²⁹,—CO—N(R²⁹)₂; —N(R²⁹)₂, —NR²⁹—CO—R′, —NR²⁹CO₂—R²⁹ wherein each R²⁹ isindependently selected from hydrogen, (C1-C6) alkyl group, (C2-C6)alkenyl group, (C2-C6) alkynyl group, (C7-C19) aralkyl group, a3-7-member-ring cyclic hydrocarbon group, a 3-7 member heterocyclicgroup, a (C6-C10) aromatic group, a 6-10 member heterocyclic aromaticgroup, each of said groups is optionally substituted; and in A8 syn/antiisomers are included, V is N or CH and U is CH, CH₂, NH or N.

In other specific embodiments A-CO— is selected from:

where variables are as defined above.

In other specific embodiments, A-CO— is selected from:

where variables are as defined above.

In other specific embodiments, A-CO— is selected from the groupconsisting of the groups listed illustrated in FIG. 3 (A15-A29) where:

XX is a substituent selected from the group consisting of —OR″, —CN,—NH₂, —N(R′)₂, halogen, —SR″, —COR′″, —COOR″, and —CON(R″)₂;

YY is halogen or —CN;

R′ is hydrogen, (C1-C6) alkyl, or (C6-C12) aryl; and

R″ and R′″ are as defined above under the definition of A groups.

In specific embodiments, XX is OH, YY is Cl, for A24 R′ is (C1-C3)alkyl,

In another embodiment, A is a benzyl group or an optionally substitutedbenzyl group. One or more moieties or groups in the A groups areprotected with one or more protecting groups.

In specific embodiments, Y is hydrogen or Y—CO— is an ester that isreadily hydrolyzed in vivo.

In another embodiment, the invention provides compounds of formula:

and pharmaceutically acceptable salts thereof, where variables are asdefined above. In specific embodiments, R⁴ and R⁵ are both hydrogens. Inspecific embodiments, R is A-CO—NH—. In specific embodiments, R isbenzyl-NH.

In another embodiment, the invention provides compounds of formula:

and pharmaceutically acceptable salts thereof, where variables are asdefined above. In specific embodiments, R⁴ and R⁵ are both hydrogens. Inspecific embodiments, R is A-CO—NH—. In specific embodiments, R isbenzyl-NH—.

In another embodiment, the invention provides compounds of formula:

and pharmaceutically acceptable salts thereof, where variables are asdefined above. In specific embodiments, R⁴ and R⁵ are both hydrogens. Inspecific embodiments, R is A-CO—NH—. In specific embodiments, R isbenzyl-NH—.

In another embodiment, the invention provides compounds of formula:

and pharmaceutically acceptable salts thereof, where variables are asdefined above. In specific embodiments, R⁴ and each R⁵ is a hydrogen. Inspecific embodiments, R is A-CO—NH—. In specific embodiments, R isbenzyl-NH—. In specific embodiments, R⁸⁻¹⁰ in M groups B, BZ, or F areelectron withdrawing groups, including esters, carbamates and alkylfunctionalized carbonyl groups.

In additional embodiments, the invention provides compounds of formulasC1-C11 as in FIG. 4. In specific embodiments of each of C1-C11, R⁴ is ahydrogen. In specific embodiments, R is A-CO—NH—. In specificembodiments, R is benzyl-NH—.

In specific embodiments, the invention provides compounds of FormulasS1, S2 and S3 (FIG. 4) where the stereochemistry of certain ringsubstituents is shown. In specific embodiments, R⁴ and each R⁵ is ahydrogen. In specific embodiments, R is A-CO—NH—. In specificembodiments, R is benzyl-NH—, where the benzyl group is optionallysubstituted. Optional substitution includes substitution with one ormore C1-C3 alkyl, C6-C12 aryl, C1-C3 haloalkyl (e.g., —CF₃), halogen, I,Cl, Br, F, —OH, or —NH₂.

In specific embodiments, the invention provides compounds of any offormulas I, II, III, IV, V, C1-C11, and S1-S3 herein wherein M isselected from one of P1-P12 (FIG. 5), where each Ha, independently, ishalogen and X and R′ of P1-P12 are as defined above. In specificembodiments, each X, independently, is a good leaving group as definedherein. In specific embodiments, each Ha, independently, is I, Br or Cl.In specific embodiments, each X is C1. In specific embodiments, each Xis Br. In specific embodiments, each X is a pyridinium group. Inspecific embodiments, one X is a pyridinium group. In specificembodiments, each R′, independently, is hydrogen, C1-C6 alkyl, or C6-C12aryl, both of which are optionally substituted. In specific embodiments,each R′, independently, is hydrogen or C1-C3 alkyl which is optionallysubstituted. In specific embodiments, each R′ is hydrogen. In specificembodiments, all X are the same. Optional substitution includessubstitution with one or more C1-C3 alkyl, C6-C12 aryl, C1-C3 haloalkyl(e.g., —CF₃), halogen, I, Cl, Br, F, —OH, or —NH₂.

In specific embodiments, the invention provides compounds of any offormulas I, II, III, IV, V, C1-C11, and S1-S3 wherein M is selected fromone of B1-B5 (FIG. 5), where X and R′ of the B1-B5 are as defined above.In specific embodiments, each X, independently, is a good leaving groupas defined herein. In specific embodiments, each X, independently, is I,Br or Cl. In specific embodiments, each X is C1. In specificembodiments, each X is Br. In specific embodiments, each X is apyridinium group. In specific embodiments, each R′, independently, ishydrogen, C1-C6 alkyl, or C6-C12 aryl, both of which are optionallysubstituted. In specific embodiments, each R′, independently, ishydrogen or C1-C3 alkyl which is optionally substituted. In specificembodiments, each R′ is hydrogen. In specific embodiments, all X are thesame. In specific embodiments, two X are the same. Optional substitutionincludes substitution with one or more C1-C3 alkyl, C6-C12 aryl, C1-C3haloalkyl (e.g., —CF₃), halogen, I, Cl, Br, F, —OH, or —NH₂.

In specific embodiments, the invention provides compounds of any offormulas I, II, III, IV, V, C1-C11, and S1-S3 wherein M is selected fromone of BZ1-BZ5 (FIG. 5), where X and R′ of the BZ1-BZ5 are as definedabove. In specific embodiments, each X, independently, is a good leavinggroup as defined herein. In specific embodiments, each X, independently,is I, Br or Cl. In specific embodiments, each X is Cl. In specificembodiments, each X is Br. In specific embodiments, each X is apyridinium group. In specific embodiments, each R′, independently, ishydrogen, C1-C6 alkyl, or C6-C12 aryl, both of which are optionallysubstituted. In specific embodiments, each R′, independently, ishydrogen or C1-C3 alkyl which is optionally substituted. In specificembodiments, each R′ is hydrogen. In specific embodiments, all X are thesame. In specific embodiments, two X are the same. Optional substitutionincludes substitution with one or more C1-C3 alkyl, C6-C12 aryl, C1-C3haloalkyl (e.g., —CF₃), halogen, I, Cl, Br, F, —OH, or —NH₂.

In specific embodiments, the invention provides compounds of any offormulas I, II, III, IV, V, C1-C11, and S1-S3 wherein M is selected fromone of D1-D3 (FIG. 5), where R′ of D1-D3 is as defined above. Inspecific embodiments, each R′, independently, is hydrogen, C1-C6 alkyl,or C6-C12 aryl, both of which are optionally substituted. In specificembodiments, each R′, independently, is hydrogen or C1-C3 alkyl which isoptionally substituted. In specific embodiments, each R′ is hydrogen. Inspecific embodiments, all R′ are the same.

In specific embodiments, the invention provides compounds of any offormulas I, II, III, IV, V, C1-C11, and S1-S3 wherein M is selected fromone of E1-E8 (FIG. 5), where X and R′ of E1-E8 are as defined above. Inspecific embodiments, each X, independently, is a good leaving group asdefined herein. In specific embodiments, each X, independently, is I, Bror Cl. In specific embodiments, each X is C1. In specific embodiments,each X is Br. In specific embodiments, each X is a pyridinium group. Inspecific embodiments, each R′, independently, is hydrogen, C1-C6 alkyl,or C6-C12 aryl, both of which are optionally substituted. In specificembodiments, each R′, independently, is hydrogen or C1-C3 alkyl which isoptionally substituted. In specific embodiments, each R′ is hydrogen. Inspecific embodiments, all X are the same. In specific embodiments, two Xare the same. In specific embodiments, X directly bonded to the ring isdifferent from X indirectly bonded to the ring. In specific embodiments,X directly bonded to the ring is a halogen and the X indirectly bondedto the ring is not a halogen. Optional substitution includessubstitution with one or more C1-C3 alkyl, C6-C12 aryl, C1-C3 haloalkyl(e.g., —CF₃), halogen, I, Cl, Br, F, —OH, or —NH₂.

In specific embodiments, the invention provides compounds of any offormulas I, II, III, IV, V, C1-C11, and S1-S3 wherein M is selected fromone of F1-F8 (FIG. 5), where X and R′ of F1-F8 are as defined above. Inspecific embodiments, each X, independently, is a good leaving group asdefined herein. In specific embodiments, each X, independently, is I, Bror Cl. In specific embodiments, each X is Cl. In specific embodiments,each X is Br. In specific embodiments, each X is a pyridinium group. Inspecific embodiments, one X is a pyridinium group. In specificembodiments, each R′, independently, is hydrogen, C1-C6 alkyl, or C6-C12aryl, both of which are optionally substituted. In specific embodiments,each R′, independently, is hydrogen or C1-C3 alkyl which is optionallysubstituted. In specific embodiments, each R′ is hydrogen. In specificembodiments, all X are the same. In specific embodiments, two X are thesame. In specific embodiments, X directly bonded to the ring isdifferent from X indirectly bonded to the ring. In specific embodiments,X directly bonded to the ring is a halogen and the X indirectly bondedto the ring is not a halogen. Preferred substituents for optionalsubstitution includes among others substitution with one or more C1-C3alkyl, C6-C12 aryl, C1-C3 haloalkyl (e.g., —CF₃), halogen, I, Cl, Br, F,—CN, —OH, C1-C3 alkoxy, —O-aryl, —O— benzyl, -phenoxy, —SH, —SR (where Ris C1-C3 alky, benzyl or phenyl), —NH₂, —N(R)₂ (where each R is C1-C3alkyl, benzyl or phenyl). Alkyl, aryl, benzyl, phenyl groups of thesesubstituents are in turn optionally substituted.

In specific embodiments, isomers of the compounds of formulas II-V,C1-C11, and S1-S3 in which the M group is cis to the R¹ group are alsoprovided.

In specific embodiments, the compounds of formula I exclude cefprozil,cefdinir, cefditoren, cefixime, and ceftobiprole. However, othercompounds of formula I maybe combined with one or more of cefprozil,cefdinir, cefditoren, cefixime, or ceftobiprole in pharmaceuticalcompositions or in medicaments.

FIG. 6 illustrates preferred stereochemistry of various core beta-lactamstructures of the formulas of this invention. In specific embodiments,compounds of the invention include those of any formula herein whichalso have the illustrated preferred stereochemistry in FIG. 6.

The term “alkyl” refers to a monoradical of a branched or unbranched(straight-chain or linear) saturated hydrocarbon and to cycloalkylgroups having one or more rings. Unless otherwise indicated preferredalkyl groups have 1 to 22 (C1-C22) carbon atoms and more preferred arethose that contain 1-12 carbon atoms (C1-12). Short alkyl groups arethose having 1 to 6 carbon atoms and those having 1-3 carbon atoms(C1-C3), including methyl, ethyl, propyl, butyl, pentyl and hexylgroups, including all isomers thereof. Long alkyl groups are thosehaving 8-30 carbon atoms and preferably those having 12-22 carbon atoms(C12-C22). The term “cycloalkyl” refers to cyclic alkyl groups havingpreferably 3 to 12 (C3-C12) carbon atoms having a single cyclic ring ormultiple condensed rings. Descriptions herein with respect to alkenylgroups apply generally to cycloalkenyl groups. Cycloalkyl groupsinclude, by way of example, single ring structures such as cyclopropyl,cyclobutyl, cyclopentyl, cyclohexyl, cyclooctyl, and the like, ormultiple ring structures such as adamantanyl, and the like. Unlessotherwise indicated alkyl groups including cycloalkyl groups areoptionally substituted as defined below.

The term “alkenyl” refers to a monoradical of a branched or unbranchedunsaturated hydrocarbon group having one or more double bonds and tocycloalkenyl groups having one or more rings wherein at least one ringcontains a double bond. Unless otherwise indicated alkyl groups have 2to 22 carbon atoms (C2-22) and more preferred are those that contain2-12 carbon atoms (C2-12). Alkenyl groups may contain one or more doublebonds (C═C) which may be conjugated or unconjugated. Preferred alkenylgroups as substituents are those having 1 or 2 double bonds and includeomega-alkenyl groups. Alkenyl groups can contain 2-5, 4, 3, or 2conjugated double bonds. Alkenyl groups include those having 2 to 6carbon atoms (C2-C6) and those having 2-3 carbon atoms (C2-C3),including ethylene (vinyl), propylene, butylene, pentylene and hexylenegroups including all isomers thereof. The term “cycloalkenyl” refers tocyclic alkenyl groups of from 3 to 22 carbon atoms (C3-C22) having asingle cyclic ring or multiple condensed rings in which at least onering contains a double bond (C═C). Descriptions herein with respect toalkenyl groups apply generally to cycloalkenyl groups. Cycloalkenylgroups preferably have 3-12 carbon atoms (C3-C12). Cycloalkenyl groupsinclude, by way of example, single ring structures (monocyclic) such ascyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cyclooctenyl,cylcooctadienyl and cyclooctatrienyl as well as multiple ringstructures. Unless otherwise indicated alkenyl groups includingcycloalkenyl groups are optionally substituted as defined below.

The term “alkynyl” refers to a monoradical of an unsaturated hydrocarbonhaving one or more triple bonds (CC). Unless otherwise indicatedpreferred alkyl groups have 2 to 22 carbon atoms and more preferred arethose that contain 2-12 carbon atoms. Alkynyl groups include ethynyl,propargyl, and the like. Short alkynyl groups are those having 2 to 6carbon atoms (C2-C6) and those including 2 or 3 carbon atoms (C2-C3),including all isomers thereof. Longer alkynyl groups are those having6-12 carbon atoms (C6-C12). The term “cycloalkynyl” refers to cyclicalkynyl groups of from 3 to 22 (C3-C22) carbon atoms having a singlecyclic ring or multiple condensed rings in which at least one ringcontains a triple bond (C≡C). Descriptions herein with respect toalkynyl groups apply generally to cycloalkynyl groups. Unless otherwiseindicated alkynyl groups including cycloalkynyl groups are optionallysubstituted as defined below.

The term “aryl” refers to a chemical group containing an unsaturatedaromatic carbocyclic group of from 6 to 22 carbon atoms (C6-C22) havinga single ring (e.g., phenyl), one or more rings (e.g., biphenyl) ormultiple condensed (fused) rings, wherein at least one ring is aromatic(e.g., naphthyl, dihydrophenanthrenyl, fluorenyl, or anthryl). An arylgroup is formally formed by removal of a hydrogen from an aryl compound.Aryls include phenyl, naphthyl and the like. Aryl groups contain maycontain portions that are alkyl, alkenyl or akynyl in addition to theunsaturated aromatic ring(s). The term “alkaryl” refers to the arylgroups containing alkyl portions, i.e., -alkylene-aryl and -substitutedalkylene-aryl. Such alkaryl groups are exemplified by benzyl, phenethyland the like.

The term “heterocyclyl” generically refers to a monoradical thatcontains at least one ring of atoms, typically a 3-10 member ring,preferably a 5, 6 or 7 member ring which may be a saturated orunsaturated ring (e.g., containing double bonds) wherein the ring cancontain one or more carbon atoms and one or more heteroatoms (anon-carbon atom). Heterocyclic groups can contain 1, 2 or 3 rings (2 ormore rings can be designated a ring system) at least one of which is aheterocyclic ring. To satisfy valence the heteroatom may be bonded to Hor a substituent group. Ring carbons may be replaced with —O—, —S—,—NR²¹—, —N═, among others, where R²¹ in this definition is hydrogen oran alkyl, aryl, heterocyclyl or heteroaryl group. Several heterocyclicgroups, rings and ring systems are more specifically described in thespecification hereof.

The term “heteroaryl” refers to a group that contains at least onearomatic ring (typically a 5 or 6-member ring) in which one or more ofthe ring carbons is replaced with a heteroatom (non-carbon atom). Tosatisfy valence the heteroatom may be bonded to H or a substituentgroup. Ring carbons may be replaced with —O—, —S—, —NR²²—, —N═, amongothers, where R²² in this definition is hydrogen or an alkyl, aryl,heterocyclyl or heteroaryl group. Heteroaryl groups may include one ormore aryl groups (all-carbon aromatic rings) or heteroaryl rings andaryl rings of the heteroaryl group may be linked by a single bond or alinker group (e.g., alkylene (CH₂)_(n)) or may be fused. Heteroarylgroups include those having aromatic rings with 5 or 6 ring atoms ofwhich 1-3 ring atoms are heteroatoms. Preferred heteroatoms are —O—,—S—, —NR— and —N═. Heteroaryl groups include those containing 5-12carbon atoms. Unless otherwise noted heteroaryl groups are optionallysubstituted as described herein.

Haloalkyl” refers to alkyl as defined herein substituted by one or morehalo groups as defined herein, which may be the same or different.Representative haloalkyl groups include, by way of example,perfluoroalkyl groups, trifluoromethyl, difluoromethyl, chloromethyl,bromomethyl, chloro-ethyl, bromo-ethyl, chloro-cyclopropyl, 2,3-dichlorocyclopropyl, and the like. Haloalkyl groups include thosehaving 1-6 (C1-C6) and 1-3 (C1-C3) carbon atoms and which contain 1, 2,3, 5, 7, 9, 11, 13 (e.g., perchloro groups), 1-6 or 1-13 halogens.Halogens include among others, chlorine, bromine, iodine and fluorine.In certain embodiments, chlorine, bromine and iodine are preferredhalogens.

The term “haloaryl” similarly refers to an aryl group as defined hereinsubstituted by one or more by one or more halo groups as defined herein,which may be the same or different. Representative haloaryl groupsinclude among others para-halophenyl, ortho-halophenyl, meta-halophenyl,and phenyl rings carrying combinations of 2-5 halogens at ortho, meta,para positions or combinations thereof. Haloaryl groups include thosehaving 6 or 12 carbon atoms (C6 or C12) which can carry 1-5 or 1-9halogens. Haloaryls include perhalogenated aryl groups. Halogens includeamong others, chlorine, bromine, iodine and fluorine. In certainembodiments, chlorine, bromine and iodine are preferred halogens.

The term alkoxy (or alkoxide) refers to a —O-alkyl group, where alkylgroups are as defined above. The term alkenoxy (alkenoxide) refers to a—O-alkenyl group where alkenyl groups are as defined above wherein thedouble bond can in certain embodiments be positioned at the carbonbonded to the oxygen. In most substituents that are alkeneoxy groups thedouble bond is preferably not positioned at the carbon bonded to theoxygen. The term alkynoxy (alkynoxide) refers to a —O— alkynyl groupwhere alkynyl groups are as defined above and wherein a triple bond ispreferably not positioned at the carbon bonded to the oxygen. The termaryloxy, refers to an —O-aryl group. The term heteroaryloxy, refers toan —O-heteroaryl group. The term heterocyclyloxy, refers to an—O-heterocyclyl group.

The term “amino” refers generically to a —N(R²³)₂ group wherein R²³ forthis definition and independently of other R²³ is hydrogen, alkyl,alkenyl, alkynyl, aryl, heterocyclyl, or heteroaryl radical as describedabove. Two of R²³ may be linked to form a ring. An “alkyl amino” grouprefers to an amino group wherein at least one R²³ is alkyl. An “arylamino” group refers to an amino group wherein at least one R²³ is aryl.

The term “amido” refers generically to an —CO—N—(R²⁴)₂ group whereinR²⁴, for this definition, independently of other R²⁴, is hydrogen,alkyl, alkenyl, alkynyl, aryl, heterocyclyl, or heteroaryl radical asdescribed above. Two of R²⁴ may be linked to form a ring. An “alkylamido” group refers to an amido group wherein at least one R²⁴ is alkyl.An “aryl amido” group refers to an amido group wherein at least one R²⁴is aryl.

The term “aminoacyl” group “refers generically to an —NR²⁵—CO—R²⁵ groupwherein, for this definition, each R²⁵ independently is hydrogen, alkyl,alkenyl, alkynyl, aryl, heterocyclyl, or heteroaryl radical as describedabove. An “alkyl aminoacyl” group refers to an aminoacyl group whereinat least one R²⁵ is alkyl. An “aryl amido” group refers to an aminoacylgroup wherein at least one R²⁵ is aryl. A variety of amino acyl groupare more specifically described in the specification hereof.

The term “imine” refers generically to an —N═CR²⁶ ₂ group or an—CR²⁶═NR²⁶ group wherein in this definition each R²⁶ independently ofother R²⁶ is hydrogen, alkyl, alkenyl, alkynyl, aryl, heterocyclyl, orheteroaryl radical as described above. Two of R²⁶ may be linked to forma ring. An “alkyl imine” group refers to an imine group wherein at leastone R²⁶ is alkyl. An “aryl imine” group refers to an imine group whereinat least one R²⁶ bis aryl. Several imine groups are more specificallydescribed in the specification hereof.

The term “sulfenyl” refers to the radical —S—R²⁷ where R²⁷, in thisdefinition, is an alkyl, alkenyl, alkynyl, aryl, heterocyclyl, orheteroaryl radical as described above. The term “sulfhydryl” refers tothe —SH group.

The term “sulfonyl” refers to the radical —SO₂—R²⁷ where R²⁷, in thisdefinition, is an alkyl, alkenyl, alkynyl, aryl, heterocyclyl, orheteroaryl radical as described above.

The term “sulfonate” refers to the radical —SO₃—R²⁸ where R²⁸, in thisdefinition, is hydrogen, alkyl, alkenyl, alkynyl, aryl, heterocyclyl, orheteroaryl radical as described above. An “alkyl sulfonate” group refersto a sulfonate group wherein R²⁸ is alkyl. An “aryl sulfonate” grouprefers to an sulfonate group wherein at least one R²⁸ is aryl. The group—SO₃H can be in the ionic form —SO₃ ⁻.

Alkyl, alkenyl, alkynyl, aryl, heteroaryl, and heterocyclic groups orthe alkyl, alkenyl, alkynyl, aryl, heteroaryl, and heterocyclic portionsof groups are optionally substituted (unless noted otherwise) asdescribed herein and may contain 1-8 non-hydrogen substituents dependentupon the number of carbon atoms in the group and the degree ofunsaturation of the group. Alkyl, alkenyl, alkynyl, aryl, heteroaryl,and heterocyclic groups may also be unsubstituted.

Optional substitution refers to substitution with one or more of thefollowing functional groups (hydrogen is not herein considered to be afunctional group):

Halogens, hydroxyl (—OH), —CN, —SH, alkyl, alkenyl, alkynyl, aryl,heterocyclyl, heteroaryl, alkoxy, alkenoxy, alkynoxy, aryloxy,heteroaryloxy, heterocyclyloxy, sulfenyl, sulfonyl, sulfonate, amine,amido, aminoacyl, imine, —COR²⁰, —COOR²⁰, —CON(R²⁰)₂, —OCOR²⁰, —OCOR²⁰,—OCN(R²⁰)₂, haloalkyl, haloalkenyl, haloaryl, —CO—C(R²⁰)₂—CO—,—NR²⁰COR²⁰, —NR²⁰COOR²⁰, —COO⁻C⁺, where each R²⁰ in this definition isselected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heterocyclyl,heteroaryl, (which in turn are optionally substituted) and C⁺ is apharmaceutically acceptable cation (of a pharmaceutically acceptablesalt).

As to any of the above groups which contain one or more substituents, itis understood, that such groups do not contain any substitution orsubstitution patterns which are sterically impractical and/orsynthetically non-feasible. In addition, the compounds of this inventioninclude all stereochemical isomers arising from the substitution ofthese compounds.

The compounds of this invention may contain one or more chiral centers.Accordingly, this invention is intended to include racemic mixtures,diasteromers, enantiomers and mixture enriched in one or moresteroisomer. The scope of the invention as described and claimedencompasses the racemic forms of the compounds as well as the individualenantiomers and non-racemic mixtures thereof.

The compounds of the present inventions form salts which are also withinthe scope of this invention. Reference to a compound of the formulas(I-V) herein is understood to include reference to salts thereof, unlessotherwise indicated. The term “salt(s)”, as employed herein, denotesacidic and/or basic salts formed with inorganic and/or organic acids andbases. In addition, when a compound contains both a basic moiety, suchas, but not limited to an amine or a pyridine ring, and an acidicmoiety, such as, but not limited to, a carboxylic acid, zwitterions(“inner salts”) may be formed and are included within the term “salt(s)”as used herein. Pharmaceutically acceptable (i.e., non-toxic,physiologically acceptable) salts are preferred, although other saltsare also useful, e.g., in isolation or purification steps which may beemployed during preparation. Salts of the compounds of the formula I maybe formed, for example, by reacting a compound of the formula I with anamount of acid or base, such as an equivalent amount, in a medium suchas one in which the salt precipitates or in an aqueous medium followedby lyophilization.

Exemplary acid addition salts include acetates (such as those formedwith acetic acid or trihaloacetic acid, for example, trifluoroaceticacid), adipates, alginates, ascorbates, aspartates, benzoates,benzenesulfonates, bisulfates, borates, butyrates, citrates,camphorates, camphorsulfonates, cyclopentanepropionates, digluconates,dodecylsulfates, ethanesulfonates, fumarates, glucoheptanoates,glycerophosphates, hemisulfates, heptanoates, hexanoates, hydrochlorides(formed with hydrochloric acid), hydrobromides (formed with hydrogenbromide), hydroiodides, 2-hydroxyethanesulfonates, lactates, maleates(formed with maleic acid), methanesulfonates (formed withmethanesulfonic acid), 2-naphthalenesulfonates, nicotinates, nitrates,oxalates, pectinates, persulfates, 3-phenylpropionates, phosphates,picrates, pivalates, propionates, salicylates, succinates, sulfates(such as those formed with sulfuric acid), sulfonates (such as thosementioned herein), tartrates, thiocyanates, toluenesulfonates such astosylates, undecanoates, and the like.

Exemplary basic salts include ammonium salts, alkali metal salts such assodium, lithium, and potassium salts, alkaline earth metal salts such ascalcium and magnesium salts, salts with organic bases (for example,organic amines) such as benzathines, dicyclohexylamines, hydrabamines[formed with N,N-bis(dehydro-abietyl)ethylenediamine],N-methyl-D-glucamines, N-methyl-D-glucamides, t-butyl amines, and saltswith amino acids such as arginine, lysine and the like. Basicnitrogen-containing groups may be quaternized with agents such as loweralkyl halides (e.g., methyl, ethyl, propyl, and butyl chlorides,bromides and iodides), dialkyl sulfates (e.g., dimethyl, diethyl,dibutyl, and diamyl sulfates), long chain halides (e.g., decyl, lauryl,myristyl and stearyl chlorides, bromides and iodides), aralkyl halides(e.g., benzyl and phenethyl bromides), and others.

Compounds of the present invention, and salts thereof, may exist intheir tautomeric form, in which hydrogen atoms are transposed to otherparts of the molecules and the chemical bonds between the atoms of themolecules are consequently rearranged. It should be understood that alltautomeric forms, insofar as they may exist, are included within theinvention.

Additionally, inventive compounds may have trans and cis isomers and maycontain one or more chiral centers, therefore exist in enantiomeric anddiastereomeric forms. The invention includes all such isomers, as wellas mixtures of cis and trans isomers, mixtures of diastereomers andracemic mixtures of enantiomers (optical isomers). When no specificmention is made of the configuration (cis, trans or R or S) of acompound (or of an asymmetric carbon), then any one of the isomers or amixture of more than one isomer is intended. The processes forpreparation can use racemates, enantiomers, or diastereomers as startingmaterials. When enantiomeric or diastereomeric products are prepared,they can be separated by conventional methods, for example, bychromatographic or fractional crystallization. The inventive compoundsmay be in the free or hydrate form.

Exemplary basic salts include ammonium salts, alkali metal salts such assodium, lithium, and potassium salts, alkaline earth metal salts such ascalcium and magnesium salts, salts with organic bases (for example,organic amines) such as benzathines, dicyclohexylamines, hydrabamines[formed with N,N-bis(dehydro-abietyl)ethylenediamine],N-methyl-D-glucamines, N-methyl-D-glucamides, t-butyl amines, and saltswith amino acids such as arginine, lysine and the like. Basicnitrogen-containing groups may be quaternized with agents such as loweralkyl halides (e.g., methyl, ethyl, propyl, and butyl chlorides,bromides and iodides), dialkyl sulfates (e.g., dimethyl, diethyl,dibutyl, and diamyl sulfates), long chain halides (e.g., decyl, lauryl,myristyl and stearyl chlorides, bromides and iodides), aralkyl halides(e.g., benzyl and phenethyl bromides), and others.

The invention also relates to prodrug forms of the compounds of formulasI-V herein. The term “prodrug” refers to an agent that is converted intothe parent drug in vivo. A prodrug is metabolized or otherwise convertedto the biologically, pharmaceutically or therapeutically active form ofthe compound. To produce a prodrug, the pharmaceutically active compoundis modified such that the active compound will be regenerated bymetabolic processes. The prodrug may be designed to alter the metabolicstability or the transport characteristics of a drug, to mask sideeffects or toxicity, to improve the flavor of a drug or to alter othercharacteristics or properties of a drug. Prodrugs may be easier toadminister than the parent drug in some situations. For example, theprodrug may be bioavailable by oral administration but the parent isnot, or the prodrug may improve solubility to allow for intravenousadministration. Knowledge of pharmacodynamic processes and drugmetabolism in vivo, allows those of ordinary skill in the art, once apharmaceutically active compound is known, can design prodrugs of thecompound. Various forms of prodrugs are well known in the art. Forexamples of such prodrug derivatives. [see: Design of Prodrugs, editedby H. Bundgaard, (Elsevier, 1985), Methods in Enzymology, Vol. 42, atpp. 309-396, edited by K. Widder, et. al. (Academic Press, 1985); ATextbook of Drug Design and Development, edited by Krosgaard-Larsen andH. Bundgaard, Chapter 5, “Design and Application of Prodrugs,” by H.Bundgaard, at pp. 113-191 (1991); H. Bundgaard, Advanced Drug DeliveryReviews, Vol. 8, p. 1-38 (1992); H. Bundgaard, et al., Journal ofPharmaceutical Sciences, Vol. 77, p. 285 (1988); and Nogrady (1985)Medicinal Chemistry A Biochemical Approach, Oxford University Press, NewYork, pages 388-392.]

Leaving groups are typically substituents which are able to leave as astable, weakly basic species. In some cases, leaving groups leave asanions, in other they leave as neutral molecules. A “good leaving group”can be recognized as being the conjugate base of a strong acid. Goodleaving groups include, among others, halogens, particularly I, Br, andCl, —CC(O)R³⁰, —SC(O)R³⁰, —OCOR³⁰, thiol (—SH), sulfenyl (—SR³⁰),phenoxy, pentafluorphenoxy, tosyl and tosyl variants includingp-fluorotosyl, p-bromotosyl, p-nitrobenzyltosyl, pentafluorotosyl, whereR³⁰ for this definition can be selected from optionally substitutedalkyl and aryl groups, specific R³⁰ include C1-C3 alkyls, particularlymethyl groups, or pyridinium groups:

where R, in this definition, represents hydrogens or 1-5 non-hydrogengroups, which include, among others, C1-C3 alkyl groups. Leaving groupsas used herein further include species like cyclopropyl groups,including substituted cyclopropyl groups, in which the bond breaking ofthe “leaving” involves ring opening. In this case, ring strain generatedupon change in hybridization and electron withdrawing properties causethe cyclopropyl ring to open.

The term beta-lactam antibiotic is used broadly herein to refer to anycompound recognized in the art as containing a beta-lactam ringstructures, including for example those ring structures illustrated inFIG. 6, and which exhibits antibiotic activity against one or moremicroorganisms, particularly bacteria. Beta-lactam antibiotics includethose described in the following references: Queener et al. Beta-lactamAntibiotics for Clinical Use 1986 (Informa Health Care); and MitsuhashiBeta-lactam Antibiotics 1981 (Japan Scientific Societies Press).

Beta-lactam compound is most generally a compound which comprises abeta-lactam ring, see exemplary rings in FIG. 4-1 through 4-2 and FIG.6. Beta-lactam compounds of interest in this invention are those whichexhibit antibiotic activity and/or inhibition of one or morebeta-lactamases and preferably those that exhibit both activities.

The term beta-lactamases is used broadly herein to refer to enzymes fromany sources which catalyze beta-lactam ring opening. Beta-lactamases (EC3.5.2.6) are enzymes most commonly produced by bacteria. Beta-lactamasescatalyze the hydrolytic ring opening of beta-lactam rings and areresponsible for conferring bacterial resistance to beta-lactamantibiotics such as penicillins, penams, penems, cephems,cephalosporins, carbacephems, cephamycins, and monobactams. Somebeta-lactamases have evolved to thermodynamic perfection whereindiffusion of beta-lactam to beta-lactamase is the rate determining step.Many different classification systems have been used to categorizebeta-lactamases including genetic and mechanistic schemes. At thesimplest level beta-lactamases can be divided up into two categories.Serine hydrolases catalyze their reactions through the use of an activesite serine that is acylated during the reaction in a Ping-Pong-Bi-Bimechanism if water is accounted as a substrate or Uni-Bi-Bi if thesolvent water molecules are ignored. Metallo beta-lactamases catalyzehydrolysis of the amide bond of the lactam ring via direct nucleophilicattack of a water molecule using one or two Zn⁺⁺ ions. This Bi-Bimechanism if water is counted or Uni-Bi mechanism if water is ignoreddoes not proceed through an acyl enzyme intermediate.

Beta-lactamase inhibitor is also used broadly herein to refer tochemical species, particularly small molecules (e.g., molecules otherthan peptides or proteins). Beta-lactamases can be inhibited by smallmolecules via reversible competitive, noncompetitive, uncompetitive, andslow tight binding mechanisms as well as irreversibleactive-site-directed and mechanism based or suicide mechanisms. Suchinhibitor molecules decrease the catalytic rate of beta-lactamasereactions or completely prevent beta-lactamases from performing anycatalysis at all. Examples of reversible competitive inhibitors includeboronic acids. Examples of active-site-directed irreversible inhibitorsinclude phosphate or phosphonate esters. Examples of mechanism basedinhibitors include clavulanic acid, sulbactam and tazobactam.

Beta-lactam compounds of interest in this invention are those whichexhibit inhibition of one or more beta-lactamases and preferably thosethat exhibit both activities and/or antibiotic activity. Beta-lactamaseinhibitors of this invention do not include clavulanic acid, sulbactamand tazobactam, however, one or more compounds of this invention can becombined with one or more of these known inhibitors in pharmaceuticalcompositions or medicaments.

Compounds of this invention can be synthesized employing methods asdescribed herein or using routine adaptations of these methods withart-known or commercially available starting materials and reagents inview of what is generally known in the art with respect to the synthesisof the various classes of known beta-lactam inhibitors and knownbeta-lactam antibiotics. For example, synthesis of starting materialsfor synthesis of compounds of the invention and also in the synthesis ofcompounds of the invention may be achieved using well-known methods andreadily available materials, such as provided in March; Larock,Comprehensive Organic Transformations (VCH Publishers, 1989); LarockComprehensive Organic Transformations: A Guide to Functional GroupPreparations, Second Edition, (John Wiley & Sons, Inc., 1999); Smith,March March's Advanced Organic Chemistry: Reactions, Mechanisms, andStructure, Sixth Edition, (John Wiley & Sons, Inc., 2007); G. I. Georg,The Organic Chemistry Of .Beta-Lactams, (VCH 1992), Page Chemistry ofBeta-Lactams (Springer, 1992); Smith, Smith Organic Synthesis, SecondEdition (McGraw-Hill Science/Engineering/Math, 2001); Bruggink A,Synthesis of [beta]-lactam Antibiotics: Chemistry, Biocatalysis &Process Integration (Springer, 2001.)

Compounds of this invention, for example those withphenylacetyl-NH-groups as the R—NH— groups in formulas herein, can beused as intermediates in the synthesis of beta-lactam compounds havingvarious aminoacyl groups at this ring position. For example,modification of the R aminoacyl groups at the 7 position (or equivalentposition) on the core ring system) can be accomplished by those ofordinary skill in the art using art-recognized techniques, startingmaterials and reagents which are available commercially or byapplication of art-known synthetic methods. Removal of the phenylacetylgroup can be accomplished, for example, by deamidation through one ofseveral methods including the use of PCI₅, penicillin amidase,cephalosporin C amidase or penicillin acylase to give the free amine atthe 7 position (or equivalent position). The amino group can then bemodified by reacting a functionalized carboxylic acid in the presence ofpenicillin amidase under acidic conditions or by activating thefunctionalized carboxylic acid with an activating agent such ascyclohexylcarbodiimide.

Mechanisms of Beta-Lactamase Inhibition

With out wishing to be bound by any particular mechanism of action ofthe compound herein, the following mechanistic discussion is provided topresent the current view of the inventors with respect to the inhibitionof beta-lactamases by compounds of this invention. It is believed thathighly reactive electrophilic or nucleophilic sites (e.g., chemicalmoieties or groups) are generated in compounds of this invention uponopening of the β-lactam ring, particularly by one or morebeta-lactamases. The species generated on lactam ring opening aregenerated from certain latent reactive moieties or groups which areconjugated to the lactam ring in the compounds of this invention. Thesesites are believed capable of covalently binding to a beta-lactamaseenzyme nucleophile or electrophile, respectively.

Schemes 1 and 2 illustrate examples of the generation of reactivenucleophiles which will react with enzyme groups such as serine,tyrosine, histidine, thiol, amines or combinations thereof. Covalentbinding of the compounds of the invention is believed to inhibit theenzyme.

Schemes 3 and 4 illustrate examples of the generation of highly reactivenucleophilic moieties upon opening of the lactam ring. These compoundswill work particularly well against the serine beta-lactamases enzymes.Because of their nonselectivity and high reactivity, they also targetthe metallo-beta-lactamases which do not proceed through a stabileacylated enzyme intermediate. These are potent nucleophiles that canreact with the abundant electrophilic centers in proteins; e.g., thecarbonyls of the amide (peptide) bonds.

Scheme 5 illustrates inhibitor compounds of this invention which aremultifunctional suicide inhibitors wherein multiple sites becomeactivated on cleavage of the beta-lactam ring and thereby becomeavailable to alkylate the beta-lactamase enzyme.

One of ordinary skill in the art will appreciate on review of Schemes1-5 above, that alternative latent reactive moieties and groups can beintroduced onto the vinyl group substituent on the various beta-lactamring systems illustrated therein which will generate electrophilic ornucleophilic sites on cleavage of the beta-lactam ring by abeta-lactamase.

The compounds of this invention are typically administered to a patientin the form of a pharmaceutical composition. Accordingly, this inventionprovides pharmaceutical compositions comprising apharmaceutically-acceptable carrier or excipient and a therapeuticallyeffective amount of a compound of formula I—V or a pharmaceuticallyacceptable salt thereof.

Any conventional carrier or excipient may be used in the pharmaceuticalcompositions of this invention. The choice of a particular carrier orexcipient, or combinations of carriers or exipients, will depend on themode of administration being used to treat a particular patient or typeof bacterial infection. The preparation of a suitable pharmaceuticalcomposition for a particular mode of administration, such as oral,topical, inhaled, or parenteral administration, is well within theknowledge of those of ordinary skill in the pharmaceutical arts.Additionally, the ingredients for such compositions arecommercially-available. For example, conventional formulations andformulations techniques are described in Remington's PharmaceuticalSciences, 17.sup.th Ed. (Mace Publishing Co., 1985) and Banker, Rhodes(Eds) Modern Pharmaceutics 4^(th) Edition (Marcel Dekker, Inc, 2002).

The pharmaceutical compositions of this invention will typically containa therapeutically effective amount of a compound of formulas I-V or apharmaceutically-acceptable salt thereof. The pharmaceuticalcompositions of this invention can contain a combined therapeuticallyeffective amount of two or more compounds of formulas I-V orpharmaceutically-acceptable salts thereof. The pharmaceuticalcompositions of this invention can contain a combined therapeuticallyeffective amount of one or more compounds of formulas I-V orpharmaceutically-acceptable salts thereof, in combination with one ormore known beta-lactam antibiotics. Typically, such pharmaceuticalcompositions will contain from about 0.01 to about 99.99%, from about0.1 to about 99.9%, from about 1% to 99%, form about 5% to about 95%,from about 10% to about 10% or from about 10% to about 50% of the activeagent(s) of this invention. One of ordinary skill in the art knows orcan readily determine therapeutically effective amounts of knownbeta-lactam antibiotics. Compounds of this invention can exhibitantibiotic activity and/or beta-lactamase inhibition. The amount orcombined therapeutically effective amount of a compound of thisinvention for antibiotic effect may be different from that forbeta-lactamase inhibition. One or ordinary skill in the art candetermine therapeutically effective amounts of the compounds of thisinvention employing art-known techniques without undue experimentation.In pharmaceutical compositions in which a beta-lactamase inhibitor ofthis invention is combined with a known beta-lactamase antibiotic or abeta-lactamase antibiotic of this invention, the therapeuticallyeffective amount typically employed will be that for achievingbeta-lactamase inhibition.

Pharmaceutical compositions of this invention include those suitable forparenteral administration, particularly intravenous administration. Suchpharmaceutical compositions typically comprise a sterile,physiologically-acceptable aqueous solution containing a therapeuticallyeffective amount or combined amount of a compound of formulas I-V orpharmaceutically-acceptable salts thereof. Physiologically-acceptableaqueous carrier solutions suitable for intravenous administration ofactive agents are well-known in the art. Such aqueous solutions includeamong others, 5% dextrose, Ringer's solutions (lactated Ringer'sinjection, lactated Ringer's plus 5% dextrose injection, acylatedRinger's injection), Normosol-M, Isolyte E and the like. Optionally,such aqueous solutions may contain a cosolvent, for example,polyethylene glycol; a chelating agent, for example, ethylenediaminetetracetic acid; a solubilizing agent, for example, a cyclodextrin; ananti-oxidant, for example, sodium metabisulphite; and the like.

The aqueous pharmaceutical compositions of this invention can belyophilized and subsequently reconstituted with a suitable carrier priorto administration. The carrier in this composition comprises, forexample, sucrose, mannitol, dextrose, dextran, lactose or a combinationthereof.

Pharmaceutical compositions of this invention include those for oraladministration in which the active ingredient is combined with a solidcarrier or excipient. Choice of carriers and excipients for oral dosageforms is within the knowledge of one of ordinary skill the art.

The pharmaceutical compositions of this invention can be packaged in aunit dosage form. This term refers to a physically discrete unitsuitable for dosing a patient, i.e., each unit containing apredetermined quantity of active agent calculated to produce the desiredtherapeutic effect either alone or in combination with one or moreadditional units. Unit dosage forms include, among others, tablets,capsules, solutions, suspensions, elixirs, syrups, cream, lotions,ointments, sprays and aerosols. For example, such unit dosage forms maybe packaged in sterile, bottles, vials, tubes, sprayers, aerosoledispensers, sealed ampoules and the like.

Compounds of the invention are useful as antibiotics. For example, thecompounds of this invention are useful for treating or preventingbacterial infections and other bacteria-related medical conditions inmammals, including humans and animals (i.e., dogs, cats, horses, cows,pigs, etc.) which are caused by microorganisms susceptible to thecompounds of this invention. This invention provides a method oftreating a bacterial infection in a mammal, the method comprisingadministering to a mammal in need of treatment, a pharmaceuticalcomposition comprising a pharmaceutically-acceptable carrier and atherapeutically effective amount or combined therapeutically effectiveamount of one or more compounds of formulas l-V, orpharmaceutically-acceptable salts thereof.

Compounds of the invention are useful as components of antibioticcompositions. For example, the compounds of this invention are useful incombination with known beta-lactam antibiotics for treating orpreventing bacterial infections and other bacteria-related medicalconditions in mammals, including humans and animals (i.e., dogs, cats,horses, cows, pigs, etc.) which are caused by microorganisms susceptibleto the compounds of this invention. This invention provides a method oftreating a bacterial infection in a mammal, the method comprisingadministering to a mammal in need of treatment, a pharmaceuticalcomposition comprising a pharmaceutically-acceptable carrier and acombined therapeutically effective amount of a known beta-lactamantibiotic, including a beta-lactam antibiotic of this invention and oneor more beta-lactamase inhibitors of formulas I-V, orpharmaceutically-acceptable salts thereof.

Compounds of this invention are useful for treating or preventinginfections caused by Gram-positive bacteria and related microorganisms.For example, the compounds of this invention are effective for treatingor preventing infections caused by certain Enterococcus spp.;Staphylococcus spp., including coagulase negative staphylococci (CNS);Streptococcus spp.; Listeria spp.; Clostridium ssp.; Bacillus spp.; andthe like. Examples of bacterial species effectively treated with thecompounds of this invention include, but are not limited to,methicillin-resistant Staphylococcus aureus (MRSA);methicillin-susceptible Staphylococcus aureus (MSSA); glycopeptideintermediate-susceptible Staphylococcus aureus (GISA);methicillin-resistant Staphylococcus epidermitis (MRSE);methicillin-sensitive Staphylococcus epidermitis (MSSE);vancomycin-sensitive Enterococcus faecalis (EFSVS); vancomycin-sensitiveEnterococcus faecium (EFMVS); penicillin-resistant Streptococcuspneumoniae (PRSP); Streptococcus pyogenes; Bacillus anthracis and thelike.

Compounds of this invention are useful for treating or preventinginfections caused by Gram-negative bacteria and related microorganisms.For example, the compounds of this invention are effective for treatingor preventing infections cause by certain Escherichia spp.; Salmonellaspp.; Neisseria spp.; Helicobacter spp.; and the like. Examples ofbacterial species effectively treated with the compounds of thisinvention include, but are not limited to Escherichia coli 0157:H7;Salmonella enterica; Salmonella typhi; Shigella dysenteriae; Yersiniapestis; Pseudomonas aeruginosa; Vibrio cholerae; Bordetalla petussis;Haemophilus influenzae; Helicobacter pylori; Helicobacter felis;Campylobacter jejuni; Neisseria gonorrhoeae; Neisseria meningitides;Brucella abortus; Bacteroides fragilis; and the like.

Compounds of this invention are also useful for treating or preventinginfections caused by bacteria not traditionally categorized by Gramstain including but not limited to Treponema pallidum; Borreliaburgdorferi; Rickettisas spp.; and the like.

Exemplary types of infections or bacteria-related medical conditionswhich can be treated or prevented with the compounds of this inventioninclude, but are not limited to, skin and skin structure infections,urinary tract infections, pneumonia, endocarditis, catheter-relatedblood stream infections, osteomyelitis, and the like. In treating suchconditions, the patient may already be infected with the microorganismto be treated, be suspected of being infected with the microorganism ormerely be susceptible to infection in which case the active agent isadministered prophylactically.

The compounds of this invention are typically administered in atherapeutically effective amount by any acceptable route ofadministration. The compounds may be administered in a single daily doseor in multiple doses per day. The treatment regimen may requireadministration over extended periods of time, for example, for severaldays or for one to six weeks or longer. The amount of active agentadministered per dose or the total amount administered will typically bedetermined by the patient's physician and will depend on such factors asthe nature and severity of the infection, the age, weight and generalhealth of the patient, the tolerance of the patient to the active agent,the microorganism(s) causing the infection, the route of administrationand the like. Typical dosage ranges for beta-lactam antibiotics are 100mg to several grams.

Additionally, the compounds of this invention are generally effectivefor inhibiting the growth of bacteria. In this embodiment, bacteria arecontacted either in vitro or in vivo with a growth-inhibiting amount ofa compound of formula I—V or pharmaceutically-acceptable salts thereof.Inhibition of bacterial growth is typically evidenced by a decrease orlack of reproduction by the bacteria and/or by lysis of the bacteria,i.e., by a decrease in colony-forming units in a given volume over agiven period of time (i.e., per hour) compared to untreated bacteria.Compounds of this invention may be bacteriostatic or bacteriocidal.Typical concentrations of beta-lactam antibiotics effective forbacterial growth inhibition rang from tenths of micrograms to tens ofmicrograms per mL.

Additionally, the compounds of this invention are generally effectivefor inhibiting beta-lactamases. In this embodiment, the beta-lactamaseis contacted in vitro or in vivo with an inhibiting amount of a compoundof formula I—V or pharmaceutically-acceptable salts thereof. Typicaleffective concentrations of beta-lactam inhibitors for inhibitingbeta-lactamases range from tenths of micrograms to tens of microgramsper mL.

The compounds of this invention can also inhibit cell wall biosynthesisin bacteria. In this embodiment, bacterial are contacted either in vitroor in vivo with a cell wall biosynthesis-inhibiting amount of a compoundof formula I or pharmaceutically-acceptable salt thereof. Typicaleffective concentrations of beta-lactam inhibitors for inhibiting cellwall biosynthesis range from tenths of micrograms to tens of microgramsper mL.

This invention additionally relates to the use of one or more compoundsof this invention in the manufacture of a medicament for treatment ofmicrobial infection, particularly bacterial infections and particularlyinfection of bacterial which exhibit resistance to one or morebeta-lactam antibiotics because of the presence of beta-lactamases. Themedicament comprises therapeutically effective amounts or combinedamounts of one or more compounds of this invention, particularly thosecompounds which exhibit microbial and/or bacterial inhibition. Morespecifically, the invention relates to the use of one or more compoundsof the formulas herein in the manufacture of a medicament for treatmentof such microbial and bacterial infections. In specific embodiments themedicament manufactured is in suitable dosage form for oral, optical,parenteral, or other form suitable form of administration as a tablet,capsule, solution, cream ointment, or other suitable dosage for. Inspecific embodiments, the medicament further comprises apharmaceutically acceptable carrier, excipient, or diluent andparticularly a carrier or diluent suitable for oral or parenteraladministration.

This invention further relates to the use of one or more compounds ofthis invention as beta-lactamase inhibitors in the manufacture of amedicament for treatment of microbial infection, particularly bacterialinfections and particularly infection of bacterial which exhibitresistance to one or more beta-lactam antibiotics because of thepresence of beta-lactamases. In this embodiment, the medicament furthercomprises a therapeutically effective amount of a beta-lactamantibiotic. More specifically, the invention relates to the use of oneor more compounds of the formulas herein in the manufacture of amedicament for treatment of such microbial and bacterial infections. Inspecific embodiments the medicament manufactured is in suitable dosageform for oral, optical, parenteral, or other form suitable form ofadministration as a tablet, capsule, solution, cream ointment, or othersuitable dosage for. In specific embodiments, the medicament furthercomprises a pharmaceutically acceptable carrier, excipient, or diluentand particularly a carrier or diluent suitable for oral or parenteraladministration.

In specific embodiments, the invention provides the use of one or morecompounds of this invention in the manufacture of a medicament fortreatment of microbial infection, particularly bacterial infections andparticularly infection of bacterial which exhibit resistance to one ormore beta-lactam antibiotics because of the presence of beta-lactamases.In specific embodiments the medicament manufactured is in an oral orparenteral dosage form such as tablet, capsule or solution. In specificembodiments, the medicament further comprises a pharmaceuticallyacceptable carrier or diluent and particularly a carrier or diluentsuitable for oral or parenteral administration.

Without wishing to be bound by any particular theory, there can bediscussion herein of beliefs or understandings of underlying principlesand mechanisms of action relating to the invention. It is recognizedthat regardless of the ultimate correctness of any mechanisticexplanation or hypothesis, an embodiment of the invention cannonetheless be operative and useful.

When a group of substituents is disclosed herein, it is understood thatall individual members of that group and all subgroups, including anyisomers, enantiomers, and diastereomers of the group members, aredisclosed separately. When a Markush group or other grouping is usedherein, all individual members of the group and all combinations andsubcombinations possible of the group members are intended to beindividually included in the disclosure. A number of specific groups ofvariable definitions have been described herein. It is intended that allcombinations and subcombinations of the specific groups of variabledefinitions are individually included in this disclosure. Specific namesof compounds are intended to be exemplary, as it is known that one ofordinary skill in the art can name the same compounds differently.

Compounds described herein may exist in one or more isomeric forms,e.g., structural or optical isomers. When a compound is described hereinsuch that a particular isomer, enantiomer or diastereomer of thecompound is not specified, for example, in a formula or in a chemicalname, that description is intended to include each isomers andenantiomer (e.g., cis/trans isomers, R/S enantiomers) of the compounddescribed individual or in any combination.

Additionally, unless otherwise specified, all isotopic variants ofcompounds disclosed herein are intended to be encompassed by thedisclosure. For example, it will be understood that any one or morehydrogens in a molecule disclosed can be replaced with deuterium ortritium. Isotopic variants of a molecule are generally useful asstandards in assays for the molecule and in chemical and biologicalresearch related to the molecule or its use. Isotopic variants,including those carrying radioisotopes, may also be useful in biologicalresearch, diagnostic assays and in therapeutics. Methods for making suchisotopic variants are known in the art.

Molecules disclosed herein may contain one or more ionizable groups[groups from which a proton can be removed (e.g., —COOH) or added (e.g.,amines) or which can be quaternized (e.g., amines)]. All possible ionicforms of such molecules and salts thereof are intended to be includedindividually in the disclosure herein. With regard to salts of thecompounds herein, one of ordinary skill in the art can select from amonga wide variety of available counterions those that are appropriate forpreparation of salts of this invention for a given application. Inspecific applications, the selection of a given anion or cation forpreparation of a salt may result in increased or decreased solubility ofthat salt.

One of ordinary skill in the art will appreciate that synthetic methods,starting materials, reagents, beta-lactamases, beta-lactam antibiotics,commercially available beta-lactam antibiotics, enzyme assays,beta-lactamase activity assays, pharmaceutical formulations and dosageforms, other than those specifically exemplified can be employed in thepractice of the invention without resort to undue experimentation. Allart-known functional equivalents, of any such assay methods, startingmaterials, synthetic methods, starting materials, reagents,beta-lactamases, beta-lactam antibiotics, commercially availablebeta-lactam antibiotics, enzyme assays, beta-lactamase activity assays,pharmaceutical formulations and dosage forms are intended to be includedin this invention.

Whenever a range is given in the specification, for example, a range ofnumbers of elements in a chemical group or moiety (e.g., a range ofnumbers of carbons (e.g., C1-C3)), a range of any integer, a range ofany number of substituents, a temperature range, a time range, or acomposition range, all intermediate ranges and subranges, as well as allindividual values included in the ranges given are intended to beincluded in the disclosure. It will be understood that any subranges orindividual value or values in a range or subrange that are included inthe description can be excluded from the claims herein.

As used herein, “comprising” is synonymous with “including,”“containing,” or “characterized by,” and is inclusive or open-ended anddoes not exclude additional, unrecited elements or method steps. As usedherein, “consisting of” excludes any element, step, or ingredient notspecified in the claim element. As used herein, “consisting essentiallyof” does not exclude materials or steps that do not materially affectthe basic and novel characteristics of the claim. Any recitation hereinof the broad term “comprising”, particularly in a description ofcomponents of a composition or in a description of elements of a device,is intended to encompass and describe the terms “consisting essentiallyof” or “consisting of”. The invention illustratively described hereinsuitably may be practiced in the absence of any element or elements,limitation or limitations which is not specifically disclosed herein.

Although the description herein contains many specifics, these shouldnot be construed as limiting the scope of the invention, but as merelyproviding illustrations of some of the embodiments of the invention. Allreferences cited herein, other than patent documents to which priorityis claimed, are hereby incorporated by reference to the extent thatthere is no inconsistency with the disclosure of this specification.Some references provided herein are incorporated by reference herein toprovide details concerning additional starting materials, additionalmethods of synthesis, additional methods of analysis and additional usesof the invention.

The terms and expressions which have been employed are used as terms ofdescription and not of limitation, and there is no intention that in theuse of such terms and expressions of excluding any equivalents of thefeatures shown and described or portions thereof, but it is recognizedthat various modifications are possible within the scope of theinvention claimed. Thus, it should be understood that although thepresent invention has been specifically disclosed by examples, preferredembodiments and optional features, modification and variation of theconcepts herein disclosed may be resorted to by those skilled in theart, and that such modifications and variations are considered to bewithin the scope of this invention as defined by the appended claims.

All patents and publications mentioned in the specification areindicative of the levels of skill of those skilled in the art to whichthe invention pertains. References cited herein are incorporated byreference herein in their entirety to indicate the state of the art asof their publication or filing date and it is intended that thisinformation can be employed herein, if needed, to exclude specificembodiments that are in the prior art. For example, when compounds areclaimed, it should be understood that compounds known and available inthe art prior to Applicant's invention, including compounds for which anenabling disclosure is provided in the references cited herein, can beexcluded included in the compound claims herein. Some referencesprovided herein are incorporated by reference to provide detailsconcerning synthetic methods, starting materials, reagents,known-beta-lactam antibiotics, pharmaceutical formulations andcomponents of such formulations, methods of administration of suchpharmaceutical composition, purification methods, and methods ofanalysis; as well as additional uses of the invention.

THE EXAMPLES Example 1

Assay (I) for Beta-Lactamase Activity

A chromogenic cephalosporin, Cefesone, is synthesized and isolated asdescribed by Sutton et al. and used to monitor p-lactamase activity. Atypical assay monitors the hydrolysis of Cefesone via the formation of aspecies which absorbs at 486 nm (molar absorptivity constant 16,000).Absorption is monitored as a function of time in 0.1 M, pH 7.0 sodiumphosphate, 0.2 mM Cefesone and 4 volume percent DMSO cosolvent at 30° C.using a Beckman DU-40 spectrophotometer having a circulating water bathattached to the cuvette holder. The assay is initiated by addition andmixing of an appropriate amount of beta-lactamase.

Assay (II) for Beta-Lactamase Activity

Another method of monitoring for beta-lactamase activity involveddissolving enough Cefesone in ethyl acetate to make a solution of 3micrograms per microliter. Ten microliters of this solution is thenapplied to a 6 mm diffusion disc. To monitor activity, the disc isdampened with water and a small aliquot of a beta-lactamase containingsolution is applied to the disc and a color change from light yellow todeep magenta is monitored visually. Typically time is recorded to firstdetectable visible color change.

Example 2

The following example is directed to synthesis of compounds of onepreferred subset of compounds of formula I, those having a cephemnucleus and an M group having a cyclopropane ring (XX):

where variables are as defined in various formulas above. The methodapplies more specifically to compounds of formula XX where R is R′—NH—,an amine, where in formula XX, R′ most generally R is a proton or apharmacologically acceptable functional group or salt, each R¹, R², eachR″, R⁶ and R⁷, independently, are selected from hydrogens, halogens ororganic functional groups, including alkyl functionalized carbonyl,esters, carbamates, and other electron withdrawing groups. The methodmore specifically applies to compounds of formula XX where each R″, R⁶,and R⁷ are selected from the group consisting of hydrogen, halogens,carbonyl groups, alkylcarbonyl groups, alkoxycarbonyl groups, aromaticcarbonyl groups, carboxylate esters, aromatic carboxylic esters,primary, secondary, and tertiary aliphatic and aromatic amines. In thissubset of compounds the N of the beta-lactam ring system is conjugatedto the electron withdrawing cyclopropyl group in M via a pi-electronsystem. This conjugation facilitates electronic rearrangement to openthe cyclopropyl ring after the lactam ring is opened (for example by abeta-lactamase enzyme).

One method of synthesizing cephem compounds (formula XX) is by reactinga compound of formula XXI or formula XXII or reactive derivativesthereof with a compound of formula XXIII or a reactive derivativethereof:

where XX, in formulas XXI and XXII, represents any halogen such aschloride, bromide or iodide and the 4-carboxylate group (—CO—Y) can beprotected if needed to carry out the reaction;

Compounds of formula XX can also be synthesized by reacting a compoundof formula XXIV or a reactive derivative thereof with a compound offormula XXV:

After reaction, removal of any protecting groups of the 4-carboxylatecan be accomplished by conventional art-known methods. When R is anamine or functionalizing the 7-amino position with groups such as acylgroups to form aminoacyl groups can be accomplished according toconventional methods by those of ordinary skill in the art. When R is anaminoacyl group, methods for converting one aminoacyl group into anotheraminoacyl group can also be accomplished according to conventionalmethods by those of ordinary skill in the art. Isomerization of theunsaturated bonds formed in synthesis of compounds of structure XX canbe performed by conventional methods by those skilled of ordinary skillin the art. The method illustrated can be readily adapted by one ofordinary skill in the art to obtain R¹ and R² groups other thanhydrogen.

Representative Synthetic Example: Synthesis of3-Vinylcyclopropane-7-(2-Phenylacetamido)-3-Cephem-4-carboxylic acid

In 20 ml of methylene chloride and 10 ml of THF was dissolved 1 gram (2mmol) of 4-methoxybenzyl3-chloromethyl-7-(2-phenylacetamido)-3-cephem-4-carboxylate, 800 mg (3mmol) triphenylphosphine, and 1.4 grams (20 mmol) cyclopropanecarboxaldehyde. To this solution was added 400 mg (2.5 mmol) KI and 5 ml10% sodium bicarbonate. The mixture was stirred vigorously in the darkovernight according to the method of U.S. Pat. No. 6,417,351 (Jul. 9,2002) Kameyama. The aqueous phase was separated and discarded. Theorganic phase was washed thrice with water, dried with magnesiumsulfate, and concentrated. The product was purified by flash vacuumchromatography by elution first with methylene chloride which eluted theexcess triphenylphosphine and cyclopropane carboxaldehyde followed bychloroform which eluted the desired product. The fractions with similarproduct Rf on silica gel TLC with toluene to ethyl acetate (5:1 v:v)were pooled and solvent evaporated to obtain the 4-carboxyl protectedcompound (1) where stereochemistry of the cephem ring is notspecifically shown, but is that shown in formula XX:

The protected product 1 was dissolved in methylene chloride and treatedwith TFA and anisole according to the method of Lee et al. (2005) J.Organ. Chem. 70(1): 367-369. The solvents were rapidly evaporated and3-vinylcyclopropane-7-(2-phenylacetamido)-3-Cephem-4-carboxylic acid 2(again stereochemistry of the cephem ring is not shown, but is that offormula XX) was isolated by trituration with petroleum either as ayellow solid.

The methods illustrated can be employed to modify beta-lactam molecules,conferring on them the property of forming one or more reactiveintermediate in the beta-lactam compound upon opening of the lactam ringsystem. The reactive intermediate is then able to react with andirreversibly inhibit one or more beta-lactamases.

Example 3 Product Inhibition of Beta-Lactamase

The beta-lactamase assay using enzyme from Enterobacter cloacae(Sigma-Aldrich, St. Louis, Mo.) was carried out as in Example 1 (assayI) above with compound 2. After the initial rate was determined thereaction was allowed to continue for two hours. Theoretical maximalabsorbance at 486 nm is approximately 1.6 while the observed finalabsorbance was consistently 0.26. This result is consistent with productinhibition as illustrated by the following equation:

where E is enzyme, S is substrate, P is product, ES is theenzyme-substrate complex, EP is the enzyme product complex, and k₁, k⁻¹,k₂, and k⁻² are rate constants and kcat is the catalytic rate constant.In the case of product inhibition k⁻² is large with respect to k₂, so asenzyme product accumulates, more enzyme is tied up in the enzyme-productcomplex and is unavailable for catalysis.

Example 4 Time Dependent Inhibition of Beta-Lactamase by3-Vinylcyclopropane-7-(2-Phenylacetamido)-3-Cephem-4-carboxylic acid (2)

The initial control rate without inhibitor is determined by adding toone microL of 0.1 M sodium phosphate buffer (pH 7.0), 10 microL DMSO and5 microL 0.1 unit/mL beta-lactamase from Enterobacter cloacae at 30° C.The reaction is initiated by addition of 20 microL 0.1 mM Cefesone inDMSO and the initial rate is determined. Inhibition reactions aredetermined by substituting the DMSO with 0.1 mM3-vinylcyclopropane-7-(2-phenylacetamido)-3-Cephem-4-carboxylic acid 2in DMSO and incubating for increasing periods of time before initiatingthe reaction by addition of Cefesone. The inactivation profile at 200microM 3vinyl-cyclopropane-7-(2-Phenylacetamido)-3-Cephem-4-carboxylicacid is illustrated in FIG. 1. Dialysis of 10 mL 0.1 units/mLbeta-lactamase with and without 500 microM inhibitor against I L of 0.1M sodium phosphate buffer (pH 7.0) with one exchange revealed no returnof activity.

Example 5

The following example is directed to synthesis of compounds of onepreferred subset of compounds of formula I, those having a cephemnucleus and an M group having a phenyl ring carrying one or moreappropriately positioned leaving groups (XXV1). The phenyl ring and itsleaving groups are conjugated through a pi-electron system to the N ofthe beta-lactam ring.

In formula XXVI, most generally the variables are defined as for formulaI above. More specifically, R¹⁰ and R⁸ are exemplified by hydrogen,halogens, thiol, groups carrying carbonyls, alkylcarbonyl groups,alkoxycarbonyl groups, aromatic groups, substituted aromatic groups,carboxylate esters, aromatic carboxylic esters, primary, secondary, anttertiary aliphatic and aromatic amines, wherein XX represents a goodleaving group conjugated to the lactam nitrogen. More specifically, R, Yand R¹ and R² are as defined for formula XX in Example 2. Again the—CO—Y group may be a protected carbonyl, if desirable. In specificembodiments, R¹⁰ and R⁸ are hydrogens or methyl groups. In specificmethods the leaving group is a halogen, particularly I, Br or Cl,pyridinium, or thiol group, and most specifically Br.

Methods of synthesis are analogous to those in Example 2. One method ofsynthesizing cephem compounds (XXVI) is by reacting a compound of theformula XXI or Xxii (above, Example 2) or reactive derivatives thereofwith a compound of formula XXVII or a reactive derivative thereof:

Compounds of formula XXVI can also be synthesized by reacting a compoundof formula XXIV (Example 2) or a reactive derivative thereof with acompound of formula XXVIII:

where the each XX variable is the same leaving group, e. g, Br. One ofordinary skill in the art can adapt the method, if desired, so that theleaving group of the product XXVI is different from that of theintermediate XXVIII.

After reaction, removal of any protecting groups of the 4-carboxylatecan be accomplished by conventional art-known methods. When R is anamine or functionalizing the 7-amino position with groups such as acylgroups to form aminoacyl groups can be accomplished according toconventional methods by those of ordinary skill in the art. When R is anaminoacyl group methods for converting one aminoacyl group into anotheraminoacyl group can also be accomplished according to conventionalmethods by those of ordinary skill in the art. Isomerization of theunsaturated bonds formed in synthesis of compounds of structure XXVI canbe performed by conventional methods by those skilled of ordinary skillin the art. The method illustrated can be readily adapted by one ofordinary skill in the art to obtain R¹ and R² groups other thanhydrogen.

Representative Synthetic Example:Synthesis of3-(1-Bromomethyl-4-Vinylbenzene)-7-(2-Phenylacetamido)-3-Cephem-4-carboxylicacid

(1) In 75 mL acetone was dissolved 972 mg 4-methoxybenzyl3-chloromethyl-7-(2-phenylacetamido)-3-cephem-4-carboxylate (2 mmol) and410 mg KI. Immediately a fine cloudy precipitate formed. The mixture wasstirred for 3 hours, the acetone evaporated and the mixture taken up in35 mL methylene chloride. The mixture was filtered, solids discarded andmother liquor retained with the product, 4-methoxybenzyl3-iodomethyl-7-(2-phenylacetamido)-3-cephem-4-carboxylate.

(2) Triphenylphosphine (0.81 grams) was dissolved in the mother liquorand the solution stirred in the dark overnight to form 4-methoxybenzyl3-phosphonium bromidemethyl-7-(2-phenylacetamido)-3-cephem-4-carboxylate salt.

(3) The solution was diluted with an additional 50 mL methylene chloridethen vigorously stirred with 50 mL saturated sodium bicarbonate. To thisslurry was slowly added 1.22 grams 4-bromomethyl benzaldehyde dissolvedin 31 mL methylene chloride over a period of 0.5-1 hour. The reactionwas then stirred overnight.

(4) The organic layer was separated and retained, washed twice with 1.0N NaCl and dried with magnesium sulfate. The solvent was then evaporatedto give an oil residue which was then dissolved in a minimum ofmethylene chloride. The product was purified by flash vacuumchromatography by elution first with methylene chloride which eluted theexcess triphenylphosphine and cyclopropane carboxaldehyde followed bychloroform which eluted the desired product. The fractions with similarproduct Rf on silica gel TLC with toluene to ethyl acetate (5:1 v:v)were pooled and solvent evaporated to obtain the protected product 3.

The product (3) was dissolved in methylene chloride and treated with TFAand anisole according to the method of Lee et al. The solvents wererapidly evaporated and3-(1-bromomethyl-4-vinylbenzene)-7-(2-phenylacetamido)-3-Cephem-4-carboxylicacid (4) was isolated by trituration with petroleum ether as a yellowsolid.

Example 6

The assay for beta-lactamase inhibition of compound 4 is carried out asin Example 3 again using the enzyme for Enterobacter cloacae.

Time dependent inhibition of beta-lactamase by compound XI is carriedout as in Example 4. The inactivation profile at 100 microM3-(l-bromomethyl-4-vinylbenzene)-7-(2-phenylacetamido)-3-cephem-4-carboxylicacid is illustrated in FIG. 2.

Dialysis of 10 mL 0.1 units/mL beta-lactamase with and without 500microM inhibitor against 1 L of 0.1 M sodium phosphate buffer (pH 7.0)with one exchange revealed no return of activity.

Example 7 Synthesis of4-diphenylmethyl-3-[-2-(3,3-dicholoroxiran-2-yl)vinyl]-7-(2-phenylacetamido)-3-cephem-4-carboxylate

In 20 mL of methylene chloride and 10 mL of THF is dissolved 1 gram (2mmol) of4-diphenylmethyl-3-chloromethyl-7-(2-phenylacetamido)-3-cephem-4-carboxylate,800 mg (3 mmol) triphenylphosphine, and 2.8 grams (20 mmol)3,3-dichlorooxirane-2-carboxaldehyde. To this solution is added 400 mg(2.5 mmol) KI and 5 mL 10% sodium bicarbonate. The mixture is stirredvigorously in the dark overnight according to the method of Kameyamasupra. The aqueous phase is separated and discarded. The organic phaseis washed thrice with water, dried with magnesium sulfate, andconcentrated. The product is purified by flash vacuum chromatography byelution first with methylene chloride which elutes the excesstriphenylphosphine and aldehyde followed by chloroform which elutes thedesired product. The fractions with similar product Rf on silica gel TLCwith toluene to ethyl acetate (5:1 v/v) are pooled and solventevaporated to provide protected product 5.

Synthesis of3-[-2-(3,3-dicholoroxiran-2-yl)vinyl]-7-(2-phenylacetamido)-3-cephem-4-carboxylicacid

The product (5) is dissolved in methylene chloride and treated with TFAand anisole according to the method of Lee et al. The solvents arerapidly evaporated and the title compound (6) is isolated by triturationwith petroleum ether.

Example 8 Synthesis of4-diphenylmethyl-3-[-2-(2-methyl-5-oxoisoxazolidin-3-yl)vinyl]-7-(2-phenylacetamido)-3-cephem-4-carboxylate

In 20 mL of methylene chloride and 10 mL of THF is dissolved 1 gram (2mmol) of4-diphenylmethyl-3-chloromethyl-7-(2-phenylacetamido)-3-cephem-4-carboxylate,800 mg (3 mmol) triphenylphosphine, and 2.8 grams (20 mmol)3,3-dichlorooxirane-2-carboxaldehyde. To this solution is added 400 mg(2.5 mmol) KI and 5 mL 10% sodium bicarbonate. The mixture is stirredvigorously in the dark overnight according to the method of Kameyamasupra. The aqueous phase is separated and discarded. The organic phaseis washed thrice with water, dried with magnesium sulfate, andconcentrated. The product is purified by flash vacuum chromatography byelution first with methylene chloride which elutes the excesstriphenylphosphine and aldehyde followed by chloroform which elutes thetitle compound 7. The fractions with similar product Rf on silica gelTLC with 5:1 toluene to ethyl acetate is pooled and solvent evaporatedto obtain the protected product 7:

Synthesis of3-[-2-(2-methyl-5-oxoisoxazolidin-3-yl)vinyl]-7-(2-phenylacetamido)-3-cephem-4-carboxylate

The protected product 7 is dissolved in methylene chloride and treatedwith TFA and anisole according to the method of Lee et al. supra Thesolvents were rapidly evaporated and the title compound 8 is isolated bytrituration with petroleum ether.

Example 9 Synthesis of 4-methoxybenzyl-3-(1-PyridiniumMethyl-4-Vinylbenzene)-7-(2-Phenylacetamido)-3-Cephem-4-carboxylate

In 20 mL methylene chloride is dissolved 1.21 grams (2 mmol)4-methoxybenzyl-3-(1-bromomethyl-4-vinylbenzene)-7-(2-phenylacetamido)-3-cephem-4-carboxylateand 160 milligrams pyridine and stirred overnight in darkened container.The methylene chloride is removed under vacuum and the title compound 9is collected.

Synthesis of 3-(1-PyridiniumMethyl-4-Vinylbenzene)-7-(2-Phenylacetamido)-3-Cephem-4-carboxylic acid(as a bromide salt)

The protected product 9 is dissolved in methylene chloride and treatedwith TFA and anisole according to the method of Lee et aLsupra. Thesolvents are rapidly evaporated and the title compound 10 is isolated bytrituration with petroleum ether.

The foregoing examples are illustrative of the methods and compounds ofthe invention and are not intended to limit the scope of the invention.

1-19. (canceled)
 20. A method of treating bacterial infection, themethod comprising: administering to an individual in need of treatment atherapeutically effective amount of a beta-lactam antibiotic; whereinthe beta-lactam antibiotic is also a beta-lactamase inhibitor thatinhibits a beta-lactamase enzyme and which comprises a leaving groupthat is in conjugation with the lactam nitrogen of the beta-lactam ringthrough a styryl moiety such that, when the beta-lactam ring is openedby the beta-lactamase enzyme to form a ring-opened compound, the leavinggroup is expelled to form a reactive moiety on the ring-opened compound,and the reactive moiety covalently binds to the beta-lactamase enzyme,wherein the reactive moiety is a substituted or unsubstitutedcyclohexadienyl methide.
 21. The method of claim 20, wherein the styrylmoiety has the structure:

wherein: X is a leaving group; the phenyl ring is cis or trans withrespect to R¹; R¹ and R², are independently selected from the groupconsisting of hydrogen, halogen, (C1-C6) alkyl, (C1-C6) alkoxy and(C1-C6) thioalkyl; and each R⁸ and each R¹⁰ is independently selectedfrom hydrogen, halogen, (C1-C6) alkyl, (C2-C6) alkenyl, (C2-C6) alkynyl,(C7-C19) aralkyl, a 3-7-member-ring cyclic hydrocarbon, a 3-7 memberheterocyclic, a (C6-C10) aromatic, a 6-10 member heterocyclic aromatic,a (C1-C6) alkoxy, a (C1-C6) thioalkyl, a —CO—R′, —CO₂R′, —CO—N(R′)₂, and—N(F)₂, wherein each R′ is independently selected from hydrogen, (C1-C6)alkyl, (C2-C6) alkenyl, (C2-C6) alkynyl, (C7-C19) aralkyl, a3-7-member-ring cyclic hydrocarbon, a 3-7 member heterocyclic, a(C6-C10) aromatic and a 6-10 member heterocyclic aromatic group, andwherein each of said groups is optionally substituted.
 22. The method ofclaim 21, wherein R¹ and R² are both hydrogen.
 23. The method of claim21, wherein R⁸ and R¹⁰ are independently selected from hydrogen,halogen, (C1-C6) alkyl, and (C1-C6) alkoxy.
 24. The method of claim 20,wherein the styryl moiety has the structure:

wherein X is a leaving group.
 25. The method of claim 20, wherein thecyclohexadienyl methide has the structure:

wherein: R² is selected from the group consisting of hydrogen, halogen,(C1-C6) alkyl, (C1-C6) alkoxy and (C1-C6) thioalkyl; and each R⁸ and R¹⁰is independently selected from hydrogen, halogen, (C1-C6) alkyl, (C2-C6)alkenyl, (C2-C6) alkynyl, (C7-C19) aralkyl, a 3-7-member-ring cyclichydrocarbon, a 3-7 member heterocyclic, a (C6-C10) aromatic, a 6-10member heterocyclic aromatic, a (C1-C6) alkoxy, a (C1-C6) thioalkyl, a—CO—R′, —CO₂R′, —CO—N(R′)₂; and —N(R′)₂, wherein each R′ isindependently selected from hydrogen, (C1-C6) alkyl, (C2-C6) alkenyl,(C2-C6) alkynyl, (C7-C19) aralkyl, a 3-7-member-ring cyclic hydrocarbon,a 3-7 member heterocyclic, a (C6-C10) aromatic and a 6-10 memberheterocyclic aromatic group, wherein each of said groups is optionallysubstituted.
 26. The method of claim 25, wherein R² is hydrogen.
 27. Themethod of claim 25, wherein R⁸ and R¹⁰ are independently selected fromhydrogen, halogen, (C1-C6) alkyl, and (C1-C6) alkoxy.
 28. The method ofclaim 20, wherein the cyclohexadienyl methide has structure:


29. The method of claim 28, wherein the styryl moiety has the structure:

wherein X is a leaving group.
 30. The method of claim 20, wherein thebeta-lactam antibiotic is a cephalosporin, a carbapenem or a monobactam.31. The beta-lactam antibiotic of claim 20, wherein the beta-lactamantibiotic is a cephalosporin.
 32. The method of claim 20, wherein theindividual in need of treatment is an individual suffering from aninfection caused by an antibiotic resistant bacteria.
 33. The method ofclaim 20, wherein the individual in need of treatment is an individualsuffering from an infection caused by a Gram-negative bacterium.
 34. Themethod of claim 33, wherein the Gram-negative bacterium is selected fromthe group consisting of Escherichia coli 0157:H7; Salmonella enterica;Salmonella typhi; Shigella dysenteriae; Yersinia pestis; Pseudomonasaeruginosa; Vibrio cholerae; Bordetella pertussis; Haemophilusinfluenzae; Helicobacter pylori; Helicobacter felis; Campylobacterjejuni; Neisseria gonorrhoeae; Neisseria meningitides; Brucella abortus;and Bacteroides fragilis.
 35. The method of claim 20, wherein theindividual in need of treatment is an individual suffering from aninfection caused by a Gram-positive bacterium.
 36. The method of claim35, wherein the Gram-positive bacterium is a bacterium selected from thegroup consisting of: methicillin-resistant Staphylococcus aureus (MRSA),methicillin-susceptible Staphylococcus aureus (MSSA), glycopeptideintermediate-susceptible Staphylococcus aureus (GISA),methicillin-resistant Staphylococcus epidermitis (MRSE),methicillin-sensitive Staphylococcus epidermitis (MSSE),vancomycin-sensitive Enterococcus faecalis (EFSVS), vancomycin-sensitiveEnterococcus faecium (EFMVS), penicillin-resistant Streptococcuspneumoniae (PRSP), and Streptococcus pyogenes.
 37. The method of claim20, wherein said beta-lactam antibiotic also acts as a beta-lactamaseinhibitor by irreversibly binding to a beta-lactamase enzyme.
 38. Themethod of claim 20, wherein the leaving group is selected from the groupconsisting of: halide, pyridinium, thiol, —SC(O)R′, —OCOR′, sulfenyl,phenoxy, pentafluorphenoxy, and tosyl, and wherein R′ is selected fromoptionally substituted alkyl and optionally substituted aryl groups. 39.The beta-lactam antibiotic of claim 20, wherein the leaving group is ahalide or a pyridinium.
 40. The beta-lactam antibiotic of claim 20,wherein: the beta-lactam antibiotic is a cephalosporin; the styrylmoiety has the structure:

wherein X is a leaving group; and the cyclohexadienyl methide hasstructure: